Ischemia minimization is associated with improved cardiac function in an ex vivo xeno working heart model
Gannon McGrath1, Ryan Chaban1,2, Anthony Calhoun1, Peter Craffey1, Victoria Diaz1, Kohei Kinoshita1, Madelyn Ma1, Zahra Habibabady1, Pratts Shannon1, Lars Burdorf1, Jacob Layer3, Ranjith Anand3, Katherine Hall3, Michele Youd3, Kathryn Stiede3, Wenning Qin3, Mike Curtis3, Richard N Pierson III1.
1Center for Transplantation Sciences, MGH, Charlestown, MA, United States; 2Surgery, University Hospital of Mainz, Mainz, Germany; 3eGenesis Inc., Cambridge, MA, United States
Introduction Previous studies showed that ischemia minimization (IM: perfusing the heart during storage), prevents ‘initial cardiac xenograft dysfunction’ (ICXD). Here we report initial observations in a working ex vivo model of ICXD designed to evaluate effects associated with IM and genetic modifications targeted to address xeno-injury on pig heart performance during perfusion with human blood.
Method Hearts from genetically modified pigs were flushed with cold preservation solution (4oC) and then stored for 3 hours either in cold saline (4oC: cold storage (CS)) or perfused with oxygenated STEEN Solution with hRBCs (IM). IM perfusion at 30-40 mmHg was initiated at room temperature for 20 minutes before cooling to 4oC. The genetically modified hearts either had combined knockouts of three specific xenogeneic carbohydrate genes (GalKO, CMAHKO, b4GALNT2KO: TKO) with variable expression of human transgenes (hTG) including complement and thrombo-regulatory proteins (n=17); two carbohydrate genes (GalKO, b4GALNT2KO: DKO) with hCD55 (n=1); GalKO (n=3); or GalKO with hCD55 (GalKO.hCD55; n=5). Heart function and biometric parameters were assessed at specific timepoints while perfused with freshly collected heparinized human blood on a working heart perfusion apparatus. Troponin I was used as a marker for myocardial injury. Transmyocardial lactate gradient assessed cardiac metabolic status.
Results 26 hearts were perfused ex vivo, 15 after CS (TKO + hTG n=9, DKO.hCD55 n=1, GalKO n=1, GalKO.hCD55 n=4) and 11 after IM (TKO + hTG n=8, GalKO n=2, GalKO.hCD55 n=1). Cardiac output (mL/min) was significantly improved for the IM group at 5 mmHg preload relative to CS, (228 vs 165, p=0.007). No significant difference in cardiac output was detected at 10 (238 vs 204, p=0.10) or 15 (250 vs 226, p=0.29) mmHg between IM vs CS. No significant difference was observed in troponin release (60 min: 77 vs 150 ng/mL, p=0.30; 240 min: 801 vs 578, p = 0.97) or lactate extraction between IM vs CS (60 min: 390 vs 240 µmol/L, p=0.49; 240min: -240 vs -90 µmol/L, p= 0.36).
Conclusion IM is protective with respect to cardiac function at low preload conditions following pig heart ischemia and subsequent reperfusion with human blood in an ex vivo model of heart xenotransplantation. The hypothesis that variation in physiologic and biochemical outcomes within groups is influenced by variable expression of complement and coagulation pathway regulatory genes is being investigated.
